Epoxycyclohexane derivative and plant growth regulator

ABSTRACT

The present invention relates to a plant growth regulator comprising as an active ingredient an epoxycyclohexane derivative represented by general formula (1): ##STR1## wherein R 1  is a hydrogen atom, a C 1  -C 6 , alkyl group or C 3  -C 6  cycloalkyl group, and R 2  and R 3  are independently C 1  -C 6  alkyl groups or are combined to form a C 2  -C 3  polymethylene group which may be substituted with a C 1  -C 6 , alkyl group, as well as to a plant growth regulator comprising the epoxycyclohexane derivative and a brassinosteroid as active ingredients. The epoxycyclohexane derivatives exhibit a potent plant growth regulating action which is equivalent to or higher than that of abscisic acid, and are useful as plant growth regulators such as a plant growth accelerator, a germination growth accelerator, a transpiration and wilting inhibitor, a cold resistance enhancer, an accelerator for growing, thickening or maturing fruits, roots, stems or bulbs, etc.. A synergistic effect is achieved by combination of the epoxycyclohexane derivative with a brassinosteroid.

TECHNICAL FIELD

The present invention relates to novel epoxycyclohexane derivatives anda plant growth regulator with abscisic acid-like physiological actions.

BACKGROUND ART

Abscisic acid is one of plant hormones such as auxin, gibberellin,cytokinin, ethylene etc. Since abscisic acid was found in 1963, itsphysiological actions including abscission layer formation, dormancyinduction, germination suppression, flowering suppression, bolting(flower stalk development) suppression, transpiration suppression, agingpromotion, and stress resistance (e.g. cold resistance enforcement) cameto be known. Although it is assumed that abscisic acid generallyexhibits a growth suppressing action as described above, it was recentlyfound that similar to other plant hormones, abscisic acid exhibits bothpromoting and suppressing effects depending on its concentration, andfor example it promotes plant growth to raise the yield at lowconcentration (Nakabori et al., Bulletin of the Aomori AgriculturalExperiment Station in 1991 (1992)). Further application to the promotionof thickening and maturing fruits (Japanese Patent LOP Publication Nos.264,005/1992, 264,006/1992 and 264,007/1992), prevention of flowers orunmatured fruits from falling (Japanese Patent LOP Publication No.139,911/1993), growth promotion for agricultural products (JapanesePatent LOP Publication No. 178,705/1993) or flowering promotion(Japanese Patent LOP Publication No. 186,303/1993) is known.

However, abscisic acid is expensive and among optical isomers ofabscisic acid, natural type one demonstrates higher effects, and thusabscisic acid is not practically used. Recently, a method of producingnatural type abscisic acid by culturing a microorganism of the genusBotrytis was developed, but it is hard to say that this method issatisfactory (Japanese Patent LOP Publication Nos. 296,696/1988,296,697/1988 and 60,590/1990). Some reports have been made of itsorganic synthesis, but there remain problems with a large number ofsteps, costs, stereoselectivity (Helv. Chim. Acta, 71, 931 (1988); J.Org. Chem., 54, 681 (1989); and Japanese Patent LOP Publication No.184,966/1991).

Out of those compounds which relate to the plant growth regulator of thepresent invention, a free carboxylic acid and methyl ester derivativeare described in the above literatures as intermediates for chemicallysynthesizing abscisic acid, but it is not disclosed that suchintermediates exhibit abscisic acid-like physiological actions.

On one hand, brassinosteroids are a group of ubiquitous compoundspresent in plants, and exhibit specific physiological growth actions,such as growth promotion, fertilization and fructification promotion,cold resistance enforcement, promotion for thickening fruits etc., andpromotion for germinating or rooting of seeds or cuttings.

However, it was not known that an intimate mixture of abscisic acid oran abscisic acid-like substance and a brassinosteroid exhibits asynergistic effect on plant growth regulation.

DISCLOSURE OF THE INVENTION

The object of the present invention is to provide a novel and highlyactive substance showing abscisic acid-like physiological actions and ahighly active plant growth regulator.

As a result of their eager research, the present inventors, found that aspecific epoxycyclohexane derivative shows excellent abscisic acid-likephysiological actions and further that an intimate mixture of saidcompound and a brassinosteroid acts synergistically on plants to exhibita strong regulatory action on their growth, to complete the presentinvention.

That is, the present first invention relates to a plant growth regulatorcomprising as an active ingredient an epoxycyclohexane derivativerepresented by general formula (1): ##STR2## wherein R¹ is a hydrogenatom, C₁ -C₆ alkyl group or C₃ -C₆ cycloalkyl group, and R² and R³ areindependently C₁ -C₆ alkyl groups or are combined to form a C₂ -C₃polymethylene group which may be substituted with a C₁ -C₆ alkyl group,and in particular to a plant growth accelerator, a germination growthaccelerator, a transpiration and wilting inhibitor, a cold resistanceenhancer, and an accelerator for growing, thickening or maturing fruits,roots, stems or bulbs.

The present second invention relates to a plant growth regulatorcomprising as active ingredients an epoxycyclohexane derivativerepresented by general formula (1) and a brassinosteroid, and inparticular to a germination growth accelerator, a cold resistanceenhancer, and an accelerator for growing, thickening or maturing fruits,roots, stems or bulbs.

The present third invention relates to an epoxycyclohexane derivativerepresented by general formula (3): ##STR3## wherein R¹ ' is a C₂ -C₆alkyl group or C₃ -C₆ cycloalkyl group, and R² and R³ are independentlyC₁ -C₆ alkyl groups or are combined to form a C₂ -C₃ polymethylene groupwhich may be substituted with a C₁ -C₆ alkyl group.

In general formula (1), the C₁ -C₆ alkyl group represented by R¹includes a methyl group, ethyl group, propyl group, isopropyl group,butyl group, isobutyl group, s-butyl group, t-butyl group, pentyl group,isopentyl group, hexyl group, s-hexyl group etc. Among these, C₂ -C₄groups, particularly propyl and isopropyl groups, are preferred forphysiological actions.

In general formula (3), the C₂ -C₆ alkyl group represented by R¹includes an ethyl group, propyl group, isopropyl group, butyl group,isobutyl group, s-butyl group, t-butyl group, pentyl group, isopentylgroup, hexyl group, s-hexyl group etc. Among these, propyl and isopropylgroups are particularly preferred for stronger physiological actions.

In general formulae (1) and (3), the C₃ -C₆ cycloalkyl group representedby R¹ and R¹ ' includes a cyclopropyl group, cyclobutyl group,cyclopentyl group and cyclohexyl group.

In general formulae (1) and (3), the C₁ -C₆ alkyl group represented byR² and R³ is preferably a straight-chain C₁ -C₄ alkyl group and includesa methyl group, ethyl group, propyl group and butyl group. The C₂ -C₃polymethylene group which may be substituted with a C₁ -C₆ alkyl groupincludes an ethylene group, propylene group etc. Among these, anethylene group is preferred for strong activity and easy synthesis. Asubstituent group optionally present in said polymethylene groupincludes the above-described C₁ -C₆ alkyl group.

The brassinosteroids used in the present second invention includebrassinolide and its analogues ("Shokubutsu No Kagaku Chosetsu"(Chemical Regulation of Plant), 22 1!, 10-17 (1987); "Yukagaku" (OilChemistry), 39 4!, 227-235 (1990)). The analogues include compoundsdeveloped by some of the present inventors, which are represented bygeneral formula (2): ##STR4## wherein R⁴ and R⁵ are C₁ -C₆, lower alkylgroups ("Shokubutsu No Kagaku Chosetsu", 29 1!, 23-30 (1994); JapanesePatent LOP Publication No. 125,396/1989).

The C₁ -C₆, alkyl groups represented by R⁴ and R⁵ in general formula (2)are preferably C₁ -C₄ straight-chain alkyl groups and include a methylgroup, ethyl group, propyl group and butyl group. In particular, theethyl group and propyl group are preferable for high activity.

The epoxycyclohexane derivatives represented by general formula (1) or(3) are produced generally as follows: ##STR5## wherein R¹, R² and R³have the same meanings as defined above.

Epoxycyclohexanecarbaldehyde (4) as the starting material can besynthesized by a method described in a literature (Helv. Chim. Acta, 71,931 (1988)). The conversion of compound (4) into the carboxylic acid offormula (la) can be effected by the method described in Japanese PatentLOP Publication No. 184,966/1991. The present compounds represented byformula (3) can be obtained by esterifying the carboxylic acid offormula (1a), e.g. in reaction with a corresponding alcohol in thepresence of a condensation agent such as carbodiimide. Japanese PatentLOP Publication No. 184,966/1991 describes that compound (1b) of formula(1) wherein R' is a methyl group can be synthesized by allowingdiazomethane to act on carboxylic acid (1a). However, this prior methodis limited to synthesis of methyl ester and cannot be applied tosynthesis of other esters.

The compounds of formula (2) used in the present second invention areobtained generally as follows: ##STR6## wherein R⁴ and R⁵ have the samemeanings as defined above.

When (22E, 24S)-24-ethyl-5α-cholesta-2,22-diene-6-one (7) (K. Mori,Agric. Biol. Chem., 44(5), 1211 (1980)) is subjected to catalytichydroxylation with a catalytic amount of osmium tetraoxide in an inertgas such as nitrogen, argon etc. in the presence of t-butylhydroperoxide or N-methylmorpholine-N-oxide, its dihydroxylation at the2 α- and ³ α-positions proceeds selectively by regulating the amounts ofthe reactants, and 2α, 3α-dihydroxy derivative (8) can be obtained inhigh yield. This dihydroxy derivative (8) is dissolved in pyridinecontaining 4-dimethylaminopyridine and reacted with a correspondingcarboxylic anhydride (e.g. propionic anhydride, butyric anhydride, etc.)to give compound (9). Then, compound (9) is dissolved in a chlorinatedorganic solvent stable to oxidation and oxidized with organic peroxide,e.g. perbenzoic acid, m-monochloroperbenzoic acid, m-monobromoperbenzoicacid, monoperphthalic acid, trifluoroperacetic acid or their sodium orpotassium salts to give the compound of formula (2).

As the plant growth regulator according to the present first invention,said epoxycyclohexane derivative can be mixed with conventionalcarriers, diluent etc. for application to plants or plant seeds in theform of e.g. liquid, powder, emulsion, wettable powder, granules etc.Conventional plant growth regulators, herbicides, fungicides andbactericides, insecticides and acaricides etc. can also be incorporatedinto it for use. Auxiliary agents such as spreader and stickers,emulsifier, wetting agent, dispersant, fixing agent, disintegrator etc.may further be added. These carriers, diluent, auxiliary agents etc. arepreferably selected to optimize the regulatory action on plant growth.

The amount of the plant growth regulator according to the firstinvention varies depending on the application method and desired action.For application by spraying, for example, its concentration ispreferably 1000 to 1 ppm, more preferably 100 to 5 ppm. For applicationby immersion of seeds etc., its concentration is preferably 1 to 0.001ppm, more preferably 0.1 to 0.01 ppm.

As the plant growth regulator according to the present second invention,said epoxycyclohexane derivative and brassinosteroid can be mixed withconventional carriers, diluent etc. for application to plants or plantseeds in the form of e.g. liquid, powder, emulsion, wettable powder,granules etc. Conventional other plant growth regulators or herbicides,fungicides and bactericides, insecticides and acaricides etc. can alsobe incorporated into it for use. Auxiliary agents such as spreader andstickers, emulsifier, wetting agent, dispersant, fixing agent anddisintegrator may further be added. These carriers, diluent, auxiliaryagents etc. are preferably selected to optimize the regulatory action onplant growth.

The amount and mixing ratio of the epoxycyclohexane derivative and thebrassinosteroid in the plant growth regulator according to the presentsecond invention vary depending on the application method and desiredaction. For application by spraying, for example, it is preferable tomix the epoxycyclohexane derivative in the range of 100 to 0.1 ppm withthe brassinosteroid in the range of 0.1 to 0.001 ppm.

The plants to which the plant growth regulators of the present first andsecond inventions are applied include, but are not limited to,vegetables such as spinach, Chinese cabbage, cucumber, eggplant,beefsteak plant, cabbage, garland chrysanthemum, leek and onion, rootvegetables such as Japanese white radish, sweet potato, beet and potato,cereals such as rice, wheat and corn, beans such as soybean, adzuki beanand peanut, industrial crop, such as sugar cane and hemp, fruits such asgrape, tangerine, persimmon, apple, tomato, melon, pear, strawberry,peach, banana, pineapple and coffee, ornamental plants such as rubbertree, phoenix and benjamin bush, and flowers such as chrysanthemum,carnation, rose, bellflower, lily and tulip.

BEST MODE FOR CARRYING OUT THE INVENTION

Hereinafter, the present invention is described in detail by referenceto Examples and Test Examples, which however are not intended to limitthe present invention.

Example 1. Synthesis of (3b) ##STR7##

To 308 mg (1.00 mmol) of 4,4-ethylenedioxy-1-4-(hydroxycarbonyl)-3-methyl-1,3-butadiene-1-yl!-1,2-oxo-2,6,6-trimethylcyclohexane(1a) (product obtained in the same manner as in Example 2 in JapanesePatent LOP Publication No. 184,966/1991) and 180 mg (224 μL, 3.00 mmol)of propyl alcohol in dry dichloromethane (1.5 mL) was added 98 mg (0.80mmol) of p-dimethylaminopyridine (DMAP), and an argon gas was bubbledinto the mixture under cooling on ice, and it was sealed in an argonatmosphere. With stirring under cooling on ice, 227 mg (1.10 mmol) ofdicyclohexyl carbodiimide in dichloromethane (10 mL) was added to itover a period of 5 minutes, and the mixture was stirred for 15 minutesunder cooling on ice and then for 3 hours at room temperature. 10 mLdiethyl ether was further added to the reaction solution in which alarge amount of white precipitates had occurred, and the precipitateswere removed by filtration. Diethyl ether was further added, and it waswashed with an aqueous 2 M hydrochloric acid/saturated sodium chloridesolution, then with an aqueous saturated sodium hydrogen carbonatesolution, and with an aqueous saturated sodium chloride solution. Then,the diethyl ether layer was separated and dried over anhydrous sodiumsulfate. The solvent was distilled off, and the resulting crude oil, 385mg, was purified by silica gel column chromatography (16 g of Wako GelC-200 ™; hexane:ethyl acetate=4:1) to give 296 mg of4,4-ethylenedioxy-1-4-(propoxycarbonyl)-3-methyl-1,3-butadiene-1-yl!-1,2-oxo-2,6,6-trimethylcyclohexane(3b) as colorless oily matter (yield: 84%).

¹ H-NMR (CDCl₃) δ (ppm): 0.96 (3H, t, J=7.4), 1.00 (3H, s), 1.22 (3H,s), 1.25 (3H, s), 1.34 (1H, dd, J=2.1, 13.6), 1.68 (2H, dd, J=6.7, 7.4),1.74 (1H, d, J=13.6), 2.01 (3H, d, J=1.3), 2.04 (1H, dd, J=2.1, 15.7),2.28 (1H, d, J=15.7), 3.81-3.97 (4H, m), 4.07 (2H, d, J=6.7), 5.71 (1H,brs), 6.27 (1H, dd, J=0.6, 16.1), 7.62 (1H, dd, J=0.7, 16.1). LRMS m/z:350 (M⁺), 291 (M⁺ --C₃ H₇ O), 264 (M⁺ --C₄ H₆ O₂). HRMS m/z: Theoretical(as C₂₀ H₃₀ O₅) 350.2092; Found 350.2103. α!_(D) ²⁰ =10.22 (c 1.8,CHCl₃).

Example 2. Synthesis of (3a) ##STR8##

The same procedure as in Example 1 was repeated except that 138 mg (3.00mmol) of ethyl alcohol was used in place of propyl alcohol, to give 264mg of 4,4-ethylenedioxy-1-4-(ethoxycarbonyl)-3-methyl-1,3-butadiene-1-yl!-1,2-oxo-2,6,6-trimethylcyclohexane(3a) (yield: 78.5 %). ¹ H-NMR (CDCl₃) δ (ppm): 1.00 (3H, s), 1.22 (3H,s), 1.25 (3H, s), 1.38 (3H, t, J=7.1), 1.34 (1H, dd, J=2.1, 13.7), 1.75(1H, d, J=13.7), 2.01 (3H, d, J=1.3), 2.05 (1H, dd, J=2.1, 15.7), 2.28(1H, d, J=15.7), 3.82-3.96 (4H, m), 4.17 (2H, q, J=7.1), 5.70 (1H, brs),6.27 (1H, dd, J=0.6, 16.0), 7.63 (1H, dd, J=0.8, 16.0). LRMS m/z: 336(M⁺). HRMS m/z: Theoretical (as C₁₉ H₂₈ O₅) 336.1935; Found 336.1913.

Example 3. Synthesis of (3c) ##STR9##

The same procedure as in Example 1 was repeated except that 180 mg (230μL) of isopropyl alcohol was used in place of propyl alcohol, to give282 mg of 4,4-ethylenedioxy-1-4-(isopropoxycarbonyl)-3-methyl-1,3-butadiene-1-yl!-1,2-oxo-2,6,6-trimethylcyclohexane(3c) (yield: 81%). ¹ H-NMR (CDCl₃) δ (ppm): 1.00 (3H, s), 1.22 (3H, s),1.25 (3H, s), 1.26 (6H, d, J=6.3), 1.34 (1H, dd, J=2.1, 13.7), 1.74 (1H,d, J=13.7), 2.00 (3H, d, J=1.3), 2.04 (1H, dd, J=2.1, 15.8), 2.28 (1H,d, J=15.8), 3.81-3.94 (4H, m), 5.06 (1H, sept, J=6.3), 5.68 (1H, brs),6.26 (1H, dd, J=0.6, 16.1), 7.61 (1H, dd, J=0.7, 16.1). LRMS m/z: 350(M⁺), 291 (M⁺ --C₃ H₇ O), 264 (M⁺ --C₄ H₆ O₂). HRMS m/z: Theoretical (asC₂₀ H₃₀ O₅) 350.2091; Found 350.2087. α!_(D) ²⁰ =13.00 (c 1.8, CHCl₃).

Example 4. Synthesis of (3d) ##STR10##

The same procedure as in Example 1 was repeated except that 227 mg (3.00mmol) of butyl alcohol was used in place of propyl alcohol, to give 306mg of 4,4-ethylenedioxy-1-4-(butoxycarbonyl)-3-methyl-1,3-butadiene-1-yl!-1,2-oxo-2,6,6-trimethylcyclohexane(3d) (yield: 84%). ¹ H-NMR (CDCl₃) δ (ppm): 0.94 (3H, t, J=7.4), 1.00(3H, s), 1.22 (3H, s), 1.25 (3H, s), 1.34 (1H, dd, J=2.1, 13.7), 1.40(2H, tq, J=7.4, 7.4), 1.64 (2H, tt, J=6.7, 7.4), 1.74 (1H, d, J=13.7),2.01 (3H, d, J=1.2), 2.04 (1H, dd, J=2.1, 15.7), 2.28 (1H, d, J=15.7),3.82-3.96 (4H, m), 4.12 (2H, t, J=6.7), 5.70 (1H, brs), 6.27 (1H, dd,J=0.5, 16.0), 7.63 (1H, dd, J=0.7, 16.0). LRMS m/z: 364 (M⁺). HRMS m/z:Theoretical (as C₂₁ H₃₂ O₅) 364.2247; Found 364.2253.

Example 5. Synthesis of (3e) ##STR11##

The same procedure as in Example 1 was repeated except that 258 mg (3.00mmol) of cyclopentyl alcohol was used in place of propyl alcohol, togive 312 mg of 4,4-ethylenedioxy-1-4-(cyclopentyloxy-carbonyl)-3-methyl-1,3-butadiene-1-yl!-1,2-oxo-2,6,6-trimethylcyclohexane(3e) (yield: 83%). ¹ H-NMR (CDCl₃) δ (ppm): 0.96 (3H, s), 1.21 (3H, s),1.25 (3H, s), 1.34 (1H, dd, J=2.1, 13.7), 1.55-1.63 (m), 1.68-1.79 (m),1.74 (1H, d, J=13.7), 1.81-1.93 (m), 2.00 (3H, d, J=1.2), 2.04 (1H, dd,J=2.1, 15.7), 2.27 (1H, d, J=15.7), 3.82-3.96 (4H, m), 5.22 (1H, m),5.67 (1H, brs), 6.26 (1H, dd, J=0.6, 16.0), 7.60 (1H, dd, J=0.6, 16.0).LRMS m/z: 376 (M⁺). HRMS m/z: Theoretical (as C₂₂ H₃₂ O₅) 376.2247;Found 376.2226.

Synthetic Example 1. Synthesis of (2a) ##STR12##

The same procedure as described in Japanese Patent LOP Publication No.125,396/1989 was carried out to give (22R, 23R, 24S)-2α,3α-dipropionyloxy-22,23-epoxy-B-homo-7-oxa-5α-stigmastane-6-one (2a)as needle crystal. m.p.: 147°-148° C. (from methanol) ¹ H-NMR (CDCl₃) δ(ppm): 0.72 (3H, s), 1.10 (3H, s), 1.18 (3H, s), 2.73 (1H, dd), 3.00(1H, dd), 4.10 (2H, m), 4.89 (1H, m), 5.38 (1H, m). FD-MS m/z: 589 (M⁺+1).

Synthetic Example 2. Synthesis of (2b) ##STR13##

The same procedure as described in Japanese Patent LOP Publication No.125,396/1989 was carried out to give (22R, 23R, 24S)-2α,3α-dibutyroyloxy-22,23-epoxy-B-homo-7-oxa-5α-stigmastane-6-one (2b).State: amorphous. ¹ H-NMR (CDCl₃) δ (ppm): 0.67 (3H, s), 0.99 (3H, s),2.70 (1H, dd), 3.00 (1H, dd), 4.10 (2H, m), 4.86 (1H, m), 5.36 (1H, m).FD-MS m/z: 617 (M⁺ +1).

Test Example 1. Evaluation of Transpiration Suppressing and GrowthPromoting Action

Mangbean seeds were planted in vermiculite and grown at 22° C. undercontinuous fluorescent-lamp lighting. On the date when their primordialleaves were developed (epicotyl length, 2 cm), their primordial leavesand epicotyls were sprayed uniformly with a treatment solution of eachtest compound. The treatment solution was prepared by dissolving eachtest compound in a small amount of Etoll ^(T) ^(M) and diluting it withwater at a predetermined concentration.

Seven seedling per group were grown in the same manner as above in avessel with 100 mL water containing a liquid fertilizer (Hyponex™).

Four days after the treatment, i.e. when the elongation growth of theirepicotyl was completely finished, the transpiration in each treatmentgroup (reduction in the amount of water in each vessel) and theiraverage weight were determined and expressed in percentage based on thatof the non-treatment group (%). The results are shown in Tables 1 and 2.

                  TABLE 1                                                         ______________________________________                                        Results of Suppression of Transpiration                                                            Transpiration (%)*                                       Compound      Concentration (ppm)                                                                        10      1                                          ______________________________________                                        Compound (1a)          84.6    88.6                                           Compound (1b)          81.4    83.2                                           Compound (3a)          78.6    84.4                                           Compound (3b)          79.6    82.8                                           Compound (3c)          76.3    79.5                                           Compound (3d)          88.6    90.9                                           Compound (3e)          87.2    92.7                                           Natural type abscisic acid                                                                           82.7    87.6                                           Non-treatment          100.0   (4.92 ml/                                                                     plant/4 days)                                  ______________________________________                                         *Percentage relative to the nontreatment group as 100%.                  

                  TABLE 2                                                         ______________________________________                                        Results of Growth Promotion                                                                        Average                                                                       Weight of Plant (%)*                                     Compound      Concentration (ppm)                                                                        10     1                                           ______________________________________                                        Compound (1a)          110.4  107.7                                           Compound (1b)          114.9  109.3                                           Compound (3a)          113.0  111.0                                           Compound (3b)          115.2  112.6                                           Compound (3c)          116.5  113.8                                           Compound (3d)          106.1  103.0                                           Compound (3e)          105.3  101.2                                           Natural type abscisic acid                                                                           109.6  105.2                                           Non-treatment          100.0  (564 mg/plant)                                  ______________________________________                                         *Percentage relative to the nontreatment group as 100%.                  

As is evident from the above results, the compounds of the presentinvention indicated an activity which was almost equivalent to or higherthan that of natural type abscisic acid. In particular, compounds (3b)and (3c) indicated a 10-fold or more activity than that of natural typeabscisic acid.

Test Example 2. Evaluation of Seed Germination and Growth PromotingAction (1)

Unhulled rice seeds (variety: Nihon Bare) were immersed in water at 15°C. for 1 day and then immersed in an aqueous solution of each testcompound at a predetermined concentration for 24 hours. Fifteen seedswere planted in each pot (diameter: 10 cm) packed with vermiculite andgrown in a room under artificial conditions at a temperature of 20° to21° C. under continuous lighting at 15,000 lux. Meanwhile, a liquidfertilizer (Hyponex™) was given.

At the 4-leaf stage, 10 well-grown seedlings were picked up from eachpot (2 pots in each group, 20 seedlings in total), and the averageweight of seedlings including roots was determined and expressed inpercentage based on that of the non-treatment group. The results areshown in Table 3.

                  TABLE 3                                                         ______________________________________                                        Results of Promotion of Seed Germination and Growth                                               Average Weight                                                                of Seedlings (%)*                                         Compound     Concentration (ppm)                                                                        0.1    0.01                                         ______________________________________                                        Compound (1a)         103.9  106.4                                            Compound (1b)         102.7  108.8                                            Compound (3a)         104.5  111.2                                            Compound (3b)         108.2  114.9                                            Compound (3c)         110.2  113.4                                            Compound (3d)         102.8  105.3                                            Compound (3e)          99.1  102.0                                            Natural type abscisic acid                                                                          108.5  105.9                                            Non-treatment         100.0  (171 mg/seedling)                                ______________________________________                                         *Percentage relative to the nontreatment group as 100%.                  

As is evident from the above results, the compounds of the presentinvention indicated an activity which was almost equivalent to or higherthan that of natural type abscisic acid.

Test Example 3. Evaluation of Seed Germination and Growth PromotingAction (2)

Carrot seeds (variety: Koyo No. 2) were immersed instantaneously in asolution of a test compound in ethanol/water (50:50) at a predeterminedconcentration. Immediately after the treatment, the treated seeds wereair-dried, and on the next day, they were planted and cultivated in avinyl house at a temperature of 13° C. or more at night.

Sixty days after planting, the average weight of their roots wasdetermined and expressed in percentage based on that of thenon-treatment group. The results are shown in Table 4.

                  TABLE 4                                                         ______________________________________                                        Results of Promotion of Seed Germination and Growth                                          Average Weight of Roots (%)*                                   Compound       Concentration 0.1 ppm                                          ______________________________________                                        Compound (1a)  110.7                                                          Compound (1b)  115.0                                                          Compound (3a)  115.9                                                          Compound (3b)  117.2                                                          Compound (3c)  118.8                                                          Compound (3d)  113.1                                                          Compound (3e)  108.0                                                          Natural type abscisic acid                                                                   111.5                                                          Non-treatment  100.0                                                          ______________________________________                                         *Percentage relative to the nontreatment group as 100%                   

As is evident from the above results, the compounds of the presentinvention exhibited an activity which was almost equivalent to or higherthan that of natural type abscisic acid.

Test Example 4. Evaluation of Fruit Maturation Promoting Action

A grape variety, Kyoho, grown outdoors for 20 years was treated with thecompound (3c) of the present invention or natural type abscisic acid. Inthis treatment, each test compound was dissolved in 80% ethanol at apredetermined concentration and 5 ml solution was sprayed on eachcluster at the timing of beginning to color. Seventeen days afterspraying, the fruits were harvested and examined for their qualities.The results are shown in Table 5.

                  TABLE 5                                                         ______________________________________                                        Results of Promotion of Fruit Maturation                                               Concentration                                                                            Coloration                                                                              Sugar Degree                                                                           Acidity                                Compound (ppm)      Degree    (Brix %) (%)                                    ______________________________________                                        Compound (3c)                                                                          50         5.7       15.7     0.68                                   Natural  300        5.8       15.8     0.66                                   type     50         5.3       15.4     0.74                                   abscisic                                                                      acid                                                                          Non-treatment       4.6       14.7     0.80                                   ______________________________________                                         *Percentage relative to the nontreatment group as 100%                   

As is evident from the above results, the activity of 50 ppm compound(3c) according to the present invention was comparable to that of 300ppm natural type abscisic acid, indicating that the former compound hadabout 5-times activity as high as that of the latter.

Test Example 5. Evaluation of Root Thickening and Growth PromotingAction

Radishes (early var. Akamaru-commet) were cultivated in a field and atest compound was sprayed on it when their root thickening began. Thespray liquid was prepared as follows: Ninety-five parts by weight of asolvent consisting of 60 parts of xylene, 20 parts of isophorone and 20parts of a surfactant were mixed with 5 parts by weight of a testcompound to give an emulsion preparation. It was diluted with water at apredetermined concentration and then sprayed in an amount of 100liters/1,000 m².

Fifteen days after spraying, the average weight of their roots in eachgroup was determined and expressed in percentage based on that of thenon-treatment group. The results are shown in Table 6.

                  TABLE 6                                                         ______________________________________                                        Results of Root Thickening and Growth Promotion                                              Average Weight of Roots (%)*                                   Compound       Concentration 5 ppm                                            ______________________________________                                        Compound (1a)  109.8                                                          Compound (1b)  112.3                                                          Compound (3a)  111.2                                                          Compound (3b)  114.0                                                          Compound (3c)  115.4                                                          Compound (3d)  110.6                                                          Compound (3e)  109.0                                                          Natural type abscisic acid                                                                   108.5                                                          Non-treatment  100.0                                                          ______________________________________                                         *Percentage relative to the nontreatment group as 100%                   

As is evident from the above results, the compounds of the presentinvention exhibited an activity which was almost equivalent to or higherthan that of natural type abscisic acid.

Test Example 6. Evaluation of Cold Resistance Enhancing Action

Each test compound was sprayed on a Benjamin plant with 150 to 200leaves, cultivated in a pot in a greenhouse. The spraying liquid wasprepared as follows: Ninety-five parts by weight of a solvent consistingof 60 parts of xylene, 20 parts of isophorone and 20 parts of asurfactant were mixed with 5 parts by weight of a test compound to givean emulsion preparation. It was diluted with water at a predeterminedconcentration. The whole of leaves was sprayed and soaked uniformly withthe test solution.

From the day after spraying (early November), the plant was placed atambient temperatures in an open field. The percentage of fallen leavesafter 25 days was determined. The results are shown in Table 7.

                  TABLE 7                                                         ______________________________________                                        Results of Prevention of Fallen Leaves Due to Cold Damage                                    Percentage of Fallen Leave (%)                                 Compound       Concentration 10 ppm                                           ______________________________________                                        Compound (1a)  30.4                                                           Compound (1b)  25.3                                                           Compound (3a)  20.0                                                           Compound (3b)  18.8                                                           Compound (3c)  16.5                                                           Compound (3d)  27.0                                                           Compound (3e)  38.2                                                           Natural type abscisic acid                                                                   37.1                                                           Non-treatment  91.3                                                           ______________________________________                                    

As is evident from the above results, the compounds of the presentinvention exhibited the activity of preventing leaves from falling dueto cold damage, which was almost equivalent to or higher than that ofnatural type abscisic acid.

Test Example 7. Evaluation of Seed Germination and Growth PromotingAction (Combination with Brassinosteroid) (1)

Unhulled rice seeds (variety: Nihon Bare) were immersed in water at 15°C. for 1 day and then immersed in an aqueous solution of each testcompound at a predetermined concentration treatment with a singlecompound: 0.01 ppm compound; and treatment with a mixture: 0.01 ppmcompound (1a, 1b, 3a to 3e), or 0.01 ppm natural type abscisic acid,plus 0.01 ppm compound (2a)! for 24 hours. Fifteen seeds were planted ineach pot (diameter: 10 cm) packed with vermiculite and grown in a roomunder artificial conditions at a temperature of 20° to 21° C. undercontinuous lighting at 15,000 lux. Meanwhile, a liquid fertilizer(Hyponex™) was given.

At the 4-leaf stage, 10 well-grown seedlings were picked up from eachpot (2 pots in each group, 20 seedlings in total), and the averageweight of seedlings including roots was determined and expressed inpercentage based on that of the non-treatment group. The results areshown in Table 8.

                  TABLE 8                                                         ______________________________________                                        Results of Promotion of Seed Germination and Growth                                       Average Weight of Seedlings (%)*                                              treatment with                                                    Compound      a single compound                                                                           a mixture                                         ______________________________________                                        Compound (1a) 106.4         119.3                                             Compound (1b) 108.8         118.1                                             Compound (3a) 111.2         120.4                                             Compound (3b) 114.9         126.6                                             Compound (3c) 113.4         128.0                                             Compound (3d) 105.3         115.7                                             Compound (3e) 102.0         107.0                                             Compound (2a) 108.6         --                                                Natural type abscisic acid                                                                  105.9         116.5                                             Non-treatment 100.0         (171 mg/seedling)                                 ______________________________________                                         *Percentage relative to the nontreatment group as 100%.                  

As is evident from the above results, the treatment with a mixtureshowed a synergistic enhancing effect on germination and growth. Inparticular, the combination of compound (2a) with compound (3b) or (3c)showed a strong effect.

Test Example 8. Evaluation of Seed Germination and Growth PromotingAction (Combination with Brassinosteroid) (2)

Carrot seeds (variety: Koyo No. 2) were immersed instantaneously in asolution of a test compound in ethanol/water (50:50) at a predeterminedconcentration treatment with a single compound: 0.1 ppm Compound (1a,1b, 3a to 3e) or natural type abscisic acid or 0.01 ppm Compound (2a);and treatment with a mixture: 0.1 ppm Compound (1a, 1b, 3a to 3e), or0.1 ppm natural type abscisic acid, plus 0.01 ppm Compound (2a)!.Immediately after the treatment, the treated seeds were air-dried, andon the next day, they were planted and cultivated in a vinyl house at atemperature of 13° C. or more at night.

Sixty days after planting, the average weight of their roots wasdetermined and expressed in percentage based on that of thenon-treatment group. The results are shown in Table 9.

                  TABLE 9                                                         ______________________________________                                        Results of Promotion of Seed Germination and Growth                                         Average Weight of Roots (%)*                                                  treatment with                                                  Compound        a single compound                                                                          a mixture                                        ______________________________________                                        Compound (1a)   110.7        127.0                                            Compound (1b)   115.0        128.1                                            Compound (3a)   115.9        131.4                                            Compound (3b)   117.2        138.5                                            Compound (3c)   118.8        135.1                                            Compound (3d)   113.1        125.6                                            Compound (3e)   108.0        120.0                                            Compound (2a)   109.0        --                                               natural type abscisic acid                                                                    111.5        125.9                                            Non-treatment   100.0                                                         ______________________________________                                         *Percentage relative to the nontreatment qroup as 100%.                  

As is evident from the above results, the treatment with a mixtureshowed a synergistic effect on growth.

Test Example 9. Evaluation of Root Thickening and Growth PromotingAction (Combination with Brassinosteroid)

Radishes (early var. Akamaru-commet) were cultivated in a field and atest compound was sprayed on it when their root thickening began. Thespray liquid was prepared as follows: Ninety-five parts by weight of asolvent consisting of 60 parts of xylene, 20 parts of isophorone and 20parts of a surfactant were mixed with 5 parts by weight of a testcompound to give an emulsion preparation. It was diluted with water at apredetermined concentration treatment with a single compound: 5 ppmcompound (1a, 1b, 3a to 3e) or natural type abscisic acid, or 0.01 ppmcompound (2a); and treatment with a mixture: 5 ppm compound (1a, 1b, 3ato 3e), or 5 ppm natural type abscisic acid, plus 0.01 ppm compound(2a)! and then sprayed in an amount of 100 liters/1,000 m².

Fifteen days after spraying, the average weight of roots in each groupwas determined and expressed in percentage based on that of thenon-treatment group. The results are shown in Table 10.

                  TABLE 10                                                        ______________________________________                                        Results of Root Thickening and Growth Promotion                                             Average Weight of Roots (%)*                                                  treatment with                                                  Compound        a single compound                                                                          a mixture                                        ______________________________________                                        Compound (1a)   109.8        120.2                                            Compound (1b)   112.3        124.7                                            Compound (3a)   111.2        125.0                                            Compound (3b)   114.0        128.4                                            Compound (3c)   115.4        127.6                                            Compound (3d)   110.6        119.2                                            Compound (3e)   109.0        116.5                                            Compound (2a)   107.4        --                                               natural type abscisic acid                                                                    108.5        118.1                                            Non-treatment   100.0                                                         ______________________________________                                         * Percentage relative to the nontreatment group as 100%.                 

As is evident from the above results, the treatment with a mixtureshowed a synergistic effect on growth.

Test Example 10. Evaluation of Cold Resistance Enhancing Action(Combination with Brassinosteroid)

A test compound was sprayed on a Benjamin plant with 150 to 200 leaves,cultivated in a pot in a greenhouse. The spraying liquid was prepared asfollows: Ninety-five parts by weight of a solvent consisting of 60 partsof xylene, 20 parts of isophorone and 20 parts of a surfactant weremixed with 5 parts by weight of a test compound to give an emulsionpreparation. It was diluted with water at a predetermined concentrationtreatment with a single compound: 10 ppm compound (1a, 1b, 3a to 3e) ornatural type abscisic acid, or 0.01 ppm compound (2a); and treatmentwith a mixture: 10 ppm compound (1a, 1b, 3a to 3e), or 10 ppm naturaltype abscisic acid, plus 0.01 ppm compound (2a)!. The whole of leaveswas sprayed and soaked uniformly with the test solution.

From the day after spraying (early November), the plant was placed atambient temperatures in an open field. The percentage of fallen leavesafter 25 days was determined. The results are shown in Table 11.

                  TABLE 11                                                        ______________________________________                                        Results of Prevention of Fallen Leaves Due to Cold Damage                                   Percentage of Fallen Leave (%)                                                treatment with                                                  Compound        a single compound                                                                          a mixture                                        ______________________________________                                        Compound (1a)   30.4         22.4                                             Compound (1b)   25.3         18.0                                             Compound (3a)   20.0         17.1                                             Compound (3b)   18.8         13.7                                             Compound (3c)   16.5         11.8                                             Compound (3d)   27.0         25.9                                             Compound (3e)   38.2         26.0                                             Compound (2a)   28.6         --                                               natural type abscisic acid                                                                    37.1         20.8                                             Non-treatment   91.3                                                          ______________________________________                                    

As is evident from the above results, the treatment with a mixtureshowed a synergistic effect in preventing leaves from falling.

INDUSTRIAL APPLICABILITY

The epoxycyclohexane derivatives of the present invention exhibit potentplant growth regulating actions equivalent to or higher than those ofabscisic acid, such as plant growth promoting action, a germinationgrowth promoting action, transpiration and wilting preventing action,cold resistance enhancing (low-temperature-damage preventing) action,and plant thickening and growth promoting action, and are useful asplant growth regulators such as a plant growth accelerator, agermination growth accelerator, a transpiration wilting inhibitor, acold resistance enhancer, and an accelerator for growing, thickening ormaturing fruits, roots, stems or bulbs. They are also useful as plantgrowth regulators such as a regulator for falling unmatured fruits, abolting inhibitor, a preservative for cut flowers, a flowering inhibitoretc. which are known in the application of abscisic acid. Besides, theywill be applicable to brewing for improvement in qualities and reductionin costs in brewing beer. The epoxycyclohexane derivatives of thepresent invention can be easily synthesized and thus supplied in largeamounts as necessary.

The plant growth regulator of the present second invention, whichcomprises the epoxycyclohexane derivative and brassinosteroid as activeingredients, exerts synergistic actions on plant growth regulation, suchas germination growth promoting action, cold resistance enhancing(low-temperature-damage preventing) action, and plant thickening andgrowth promoting action, and are useful as plant growth regulators suchas a germination growth accelerator, a cold resistance enhancer, anaccelerator for growing, thickening or maturing fruits, roots, stems orbulbs, a cutting-rooting accelerator, etc.

What is claimed is:
 1. A plant growth repulating composition comprisingan effective amount of an epoxycyclohexane compound for regulating plantgrowth, said epoxycyclohexane compound represented by formula (1):##STR14## wherein R¹ is a hydrogen atom, a C₁ -C₆ alkyl group or C₃ -C₆cycloalkyl group, and R² and R³ are independently C₁ -C₆ alkyl groups orare combined to form a C₂ -C₃ polymethylene group which may besubstituted with C₁ -C₆ alkyl group.
 2. The plant growth regulatingcomposition according to claim 1, wherein R¹ is a propyl or isopropylgroup, and R² and R³ are combined to form an ethylene group.
 3. Anepoxycyclohexane compound represented by formula (3). ##STR15## whereinR¹ ' is a C₂ -C₆ alkyl group or C₃ -C₆ cycloalkyl group, and R² and R³are independently C₁ -C₆ alkyl groups or are combined to form a C₂ -C₃polymethylene group which may be substituted with a C₁ -C₆ alkyl group.4. The epoxycyclohexane compound according to claim 3, wherein R¹ ' is apropyl or isopropyl group.
 5. A method of regulating plant growthcomprising applying the composition according to claim 3 onto a plant.6. The method of claim 5, wherein the regulation is acceleration ofgermination growth.
 7. The method of claim 5, wherein the regulation isinhibition of transpiration and wilting.
 8. The method of claim 5,wherein the regulation is enhanced cold resistance.
 9. The method ofclaim 5, wherein the regulation is acceleration of growing, thickeningor maturing fruits, roots, stems or bulbs.
 10. The method of claim 5,wherein the regulation is acceleration of plant growth.